Terapía génica antitumoral mediante vacunas con células modificadas genéticamente.

RESUMEN Una de las estrategias más satisafactorias empleadas en la lucha contra el cáncer es la utilización de la terapia génica para actuar en la inmunomodulación, es decir la actuación sobre el propio sistema inmunitario del paciente para que sea éste quien reconozca al tumor como algo foráneo, que no escape a la inmunovigilancia y lograr destruirlo. En esta tesis doctoral hemos explorado bajo este concepto las vacunas por terapia génica no viral, tanto preventivas como terapéuticas, trabajando bien con antígenos aislados y genes como con células y genes. Los experimentos en modelo de melanoma murino, se han divido en tres grandes bloques. 1) En las vacunas preventivas con antígenos, se obtuvieron diferentes tasas de reducción de volumen tumoral, acompañadas de producción de inmunnoglobulinas G específicas frente al tumor tratado. Se obtuvieron los mejores resultados en modelos de vacunación con acondicionamiento tisular en los que inicialmente se acondicionaba al animal con los antígenos purificados, transfectando posteriormente con plásmidos desnudos productores del gen de mGM-CSF. 2) En las vacunas preventivas celulares, transfectando células B16 con plásmidos portadores de los genes de mGM-CSF, mIL-12, mB7.2 y combinaciones entre ellos, se obtuvo la protección total frente al tumor en los animales vacunados con 3 dosis de 2x105 células transfectdas, irradiadas, no seleccionadas. Además se demostró que la vacunación había generado memoria inmunológica, al repetir la exposición al tumor en los mismos animales un año después y volver a lograr la protección total. Adicionalmente se realizaron experimentos de vacunación exclusivamente con las células transfectadas, seleccionadas y purificadas mediante bolas magnéticas. 3) Por último, se realizaron experimentos de vacunación terapéutica, en animales portadores de tumor, siguiendo el modelo de células transfectadas. Se probaron diferentes dosis celulares y el tratamiento previo con ciclofosfamida. Los mejores resultados de inhibición tumoral y prolongación de la supervivencia se obtuvieron en las vacunas con 2 millones de células/dosis. Finalmente se realizó una primera aproximación al estudio del bloqueo del éxito de la vacunación terapéutica, analizando la población de células T reguladoras en los animales tratados. __________________________________________________________________________________________________One of the most satisfactory strategies against cancer is the use of gene therapy for immunemodulation, that is acting over the patients immune system in order to make it recognize the tumor and fight properly against it. In this thesis, we have explored the nonviral gene therpy vaccines under this concept, working with preventive as well as therapeutic models, with purified antigens and genes and also with whole cells and genes. The experiments have been performed with a murine melanoma model and been divided in three sets. 1) In preventive antigen vaccines, different rates of tumor growth inhibition were obtained, acompained by specific antitumor immunoglobulin G production. The best results were obtained with tissue conditioning, where the animals were firstly conditioned with purified tumor antigens and later the tissues were transfected in vivo with naked plasmids producing mGM-CSF. 2) In preventive cell vaccines, transfecting B16 cells with plasmids bearing mGM-CSF, mIL-12, mB7.2 genes alone or combined, total tumor protection was obtained vaccinating with 3 doses of 2x105 transfected, irradiated, non selected cells. This vaccine also showed immunological memory since a second challenge to the tumor in the same animals one year later also reached total protection. In addition, experiments vaccinating only with the selected population of transfected cells, by means of magnetic beads, were performed. 3) Finally, therapeutic vaccines were evaluated, continuing with transfected cells. Different cell doses and pretreatment with cyclophosphamide were tested. The best results of tumor growth inhibition and survival were obtained with 2 million transfected cells/dose. A first approach to the study of the blockade in therapeutic model success was performed analysing the regulatory T cell population in the treated animals

Mitochondrial DNA replacement techniques to prevent human mitochondrial diseases

Background: Mitochondrial DNA (mtDNA) diseases are a group of maternally inherited genetic disorders caused by a lack of energy production. Currently, mtDNA diseases have a poor prognosis and no known cure. The chance to have unaffected offspring with a genetic link is important for the affected families, and mitochondrial replacement techniques (MRTs) allow them to do so. MRTs consist of transferring the nuclear DNA from an oocyte with pathogenic mtDNA to an enucleated donor oocyte without pathogenic mtDNA. This paper aims to determine the efficacy, associated risks, and main ethical and legal issues related to MRTs. Methods: A bibliographic review was performed on the MEDLINE and Web of Science databases, along with searches for related clinical trials and news. Results: A total of 48 publications were included for review. Five MRT procedures were identified and their efficacy was compared. Three main risks associated with MRTs were discussed, and the ethical views and legal position of MRTs were reviewed. Conclusions: MRTs are an effective approach to minimizing the risk of transmitting mtDNA diseases, but they do not remove it entirely. Global legal regulation of MRTs is required

Multicompartmental Lipopolyplex as vehicle for antigens and genes delivery in vaccine formulations

Vector design and its characterization is an area of great interest in current vaccine research. In this article, we have formulated and characterized a multicompartmental lipopolyplex, which associates multiple liposomes and polyplexes in the same complex. These particles allow the simultaneous delivery of lipid or water-soluble antigens associated with genes to the same cell, in much higher amounts than conventional lipopolyplexes. The vector characterization and optimization were carried out using liposomes with entrapped carboxyfluorescein and adapted electrophoretic assays. Two types of lipopolyplexes (containing hydrophilic or lipophilic antigens) were employed to evaluate their interest in vaccination. The lipopolyplex loaded with an extract of water-soluble melanoma proteins proved to efficiently induce humoral response in murine melanoma model, increasing the levels of IgM and IgG. The specificity of the immune response induced by the lipopolyplex was demonstrated in mice with the lipopolyplex containing the GD3 ganglioside lipid antigen, abundant in melanoma cells. The levels of anti-GD3 IgG increased markedly without modifying the expression of humoral antibodies against other gangliosides

Pharmacogenetics in Neuroblastoma: What Can Already Be Clinically Implemented and What Is Coming Next?

Pharmacogenetics is one of the cornerstones of Personalized Precision Medicine that needs to be implemented in the routine of our patients' clinical management in order to tailor their therapies as much as possible, with the aim of maximizing efficacy and minimizing toxicity. This is of great importance, especially in pediatric cancer and even more in complex malignancies such as neuroblastoma, where the rates of therapeutic success are still below those of many other types of tumors. The studies are mainly focused on germline genetic variants and in the present review, state of the art is presented: which are the variants that have a level of evidence high enough to be implemented in the clinic, and how to distinguish them from the ones that still need validation to confirm their utility. Further aspects as relevant characteristics regarding ontogeny and future directions in the research will also be discussed

Integrated CGH/WES Analyses Advance Understanding of Aggressive Neuroblastoma Evolution: A Case Study

Neuroblastoma (NB) is the most common extra-cranial malignancy in preschool children. To portray the genetic landscape of an overly aggressive NB leading to a rapid clinical progression of the disease, tumor DNA collected pre- and post-treatment has been analyzed. Array comparative genomic hybridization (aCGH), whole-exome sequencing (WES), and pharmacogenetics approaches, respectively, have identified relevant copy number alterations (CNAs), single nucleotide variants (SNVs), and polymorphisms (SNPs) that were then combined into an integrated analysis. Spontaneously formed 3D tumoroids obtained from the recurrent mass have also been characterized. The results prove the power of combining CNAs, SNVs, and SNPs analyses to assess clonal evolution during the disease progression by evidencing multiple clones at disease onset and dynamic genomic alterations during therapy administration. The proposed molecular and cytogenetic integrated analysis empowers the disease follow-up and the prediction of tumor recurrence

Foxp3 Silencing with Antisense Oligonucleotide Improves Immunogenicity of an Adjuvanted Recombinant Vaccine against Sporothrix schenckii

Background: In recent years, there has been great interest in developing molecular adjuvants based on antisense oligonucleotides (ASOs) targeting immunosuppressor pathways with inhibitory effects on regulatory T cells (Tregs) to improve immunogenicity and vaccine efficacy. We aim to evaluate the immunostimulating effect of 2′OMe phosphorothioated Foxp3-targeted ASO in an antifungal adjuvanted recombinant vaccine. Methods: The uptake kinetics of Foxp3 ASO, its cytotoxicity and its ability to deplete Tregs were evaluated in murine splenocytes in vitro. Groups of mice were vaccinated with recombinant enolase (Eno) of Sporothix schenckii in Montanide Gel 01 adjuvant alone or in combination with either 1 µg or 8 µg of Foxp3 ASO. The titers of antigen-specific antibody in serum samples from vaccinated mice (male C57BL/6) were determined by ELISA (enzyme-linked immunosorbent assay). Cultured splenocytes from each group were activated in vitro with Eno and the levels of IFN-γ and IL-12 were also measured by ELISA. The results showed that the anti-Eno antibody titer was significantly higher upon addition of 8 µM Foxp3 ASO in the vaccine formulation compared to the standard vaccine without ASO. In vitro and in vivo experiments suggest that Foxp3 ASO enhances specific immune responses by means of Treg depletion during vaccination. Conclusion: Foxp3 ASO significantly enhances immune responses against co-delivered adjuvanted recombinant Eno vaccine and it has the potential to improve vaccine immunogenicity

Reseñas

[ES] Seguí de la Riva , Javier. Sobre dibujar y proyectar (por Javier Seguí de la Riva) pp. 4.-- Dibujar, proyectar LVI y LVII Arte y muerte I y II ( por Javier Seguí de la Riva) pp. 6.-- dibujar, proyectar LVIII El imaginario del dibujar (por Javier Seguí de la Riva) pp. 6 y 7.-- Dibujar, proyectar LVI y LVII Diagrama, diagramar I y II ( por Javier Seguí de la Riva) pp. 7.-- DIBUJAR, proyectar LVIII El imaginario del dibujar ( por Javier Seguí de la Riva) pp. 6 y 7.-- DIBUJAR, PROYECTAR LVI y LVII Diagrama, diagramar I y II (por Javier Seguí de la Riva) pp.7.-- DIBUJAR, PROYECTAR LIX, LX y LXI Escritos críticos I, II y III ( por Javier Seguí de la Riva) pp. 7 .-- Rehabilitacion del Antiguo Hospital San Juan de Dios para Biblioteca y Archivo Historico de Orihuela (por Ana Torres Barchino) pp. 8.-- Arnau Amo, Joaquín. Arquitectura. Ritos y ritmos (por María Elia Gutiérrez Mozo) pp. 8 y 9.—Cardone, Vito. Viaggiatori d’architettura in Italia. Da Brunelleschi a Charles Garnier (por Cesare Cundari) pp. 9 y 10.-- Casado de Amezúa Vázquez, Joaquín. Las casas reales de la Alhambra. Geometría y espacio. Una aproximación al proceso de formación del espacio ( por Pilar Chías Navarro)pp. 10.-- Jaén i Urban, Gaspar; Baldomá Soto, Montserrat y Carrasco Martí, Maria Antonia. One century of photography and preservation in catalonia: the service for local architectural heritage ( por Pilar Chías Navarro) pp. 10 y 11.-- Jaén i Urban, Gaspar. El paisaje urbano de Nueva York en la obra escrita de Federico García Lorca (por Concepción López González y Jorge García valldecabres) pp. 11.—Trachana, Angelique. Urbe Ludens ( por Gonzalo García-Rosales) pp. 12.-- Fernández-Palacios, Victoria; Yanguas, Ana; Jiménez Fdez-Palacios, Luz. Manuel Aires Mateus. Cuaderno de La Alhambra ( por José Mª Gentil Baldrich) pp. 12 y 13.-- Agustín, Luis; Miret, Elena; Vallespín, Aurelio. Representación del espacio arquitectónico. 2011.12 (por Jesús Aparicio Guisado) pp. 14.—Herschdorfer, Nathalie y Lada Umstätter, Thames. Le Corbusier and the Power of Photography (por víctor A. Lafuente Sánchez) pp. 14 y 15.-- Roma en el bolsillo. Cuadernos de dibujo y aprendizaje artístico en el siglo XVIII ( por Fernando Linares García) pp. 15 y 16.-- Vicens y Hualde, Ignacio. Dicho y hecho ( por Fernando Linares García) pp 16.-- Le Corbusier. El arte decorativo de hoy ( por Carlos Montes Serrano )pp. 16 y 17.—Jenkins Birkhäuser, Eric J. Drawn to Design: Analyzing Architecture Through Freehand Drawing ( por víctor A. Lafuente Sánchez) pp. 17 y 18.—Sobrino, Miguel. Monasterios. Las biografías desconocidas de los cenobios de España ( por Javier García-Gutiérrez Mosteiro) pp. 18 y 19.-- Jiménez Martín, Alfonso. Anatomía de la Catedral de Sevilla ( por Francisco Pinto Puerto) pp. 19.-- Raposo Grau, Javier Fco; Butragueño Díaz-Guerra, Belen y Paredes Maldonado, Miguel. Dibujar, analizar, proyectar (2010) Título I. Colección dibujo, proyecto y arquitectura ( por Mariasun Salgado de la Rosa) pp. 19 y 20.-- Raposo Grau, Fco Javier; Butragueño Díaz-Guerra, Belen y Paredes Maldonado, Miguel. Dibujar, analizar, proyectar (2011) Título III. Colección dibujo, proyecto y arquitectura ( por Carlos L. Marcos Alba) pp. 20-22.-- García Doménech, Sergio. Reflexiones urbanas sobre el espacio público de Alicante. Una interpretación de la ciudad y sus escenarios ( por Juan Calduch Cervera) pp. 22 y 23.—Cundari, Casere. Il rilievo architettonico. Ragioni. Fondamenti. Applicazioni ( por Antonio Álvaro Tordesillas) pp. 23 y 24.—Autores varios. Perspectiva-Prospettiva. La práctica de la perspectiva ( por José Mª Gentil Baldrich) pp. 24 y 25.-- Soler Sanz, Felipe. Trazados Reguladores en la Arquitectura ( por Jorge García Valldecabres) pp. 25.-- Jiménez Alcañiz, Cesar. Análisis de las metodologías para la recuperación patrimonial de entornos urbanos protegidos. Propuesta metodológica: desde los valores históricos a los nuevos modelos energéticos. Russafa desde el siglo XIX( Por Pablo Navarro Esteve) pp. 26.-- de Coca Leicher, José. El recinto ferial de la Casa de Campo de Madrid (1950-75)( por Esteban Herrero Cantalapiedra) pp. 27.-- La arquitectura religiosa renacentista en tierras del Maestre: la iglesia de Nuestra Señora de la Asunción de Vistabella del Maestrazgo ( por José Teodoro Garfella Rubio) pp. 27 y 28.-- Arquitectura de los balnearios en Galicia. Cuenca del Miño. 1816-1936 ( por José Antonio Franco Taboada)pp. 28 y 29.-- Barros da Rocha e Costa. Historia de la representación gráfica del Castillo de Peñíscola. Del grafito al láser ( por Pablo Navarro Esteve) pp. 29.-- Rivas López, Esteban José. El Carmen de la fundación Rodríguez-Acosta. Una indagación gráfica ( por Joaquín Casado de Amezúa) pp.30.-- Verdejo Gimeno, Pedro. Estaciones intermedias de ferrocarril. La sección “Non nata” Teruel-Alcañiz (por Jorge Girbés Pérez) pp. 30 y 31.-- Sender Contell, Marina. El Monasterio de Santa María de la Murta. Análisis arquitectónico de un Monasterio Jerónimo ( por Pablo Navarro Esteve) pp. 32.-- Iñarra Abad, Susana. El Render de Arquitectura. Análisis de la Respuesta Emocional del Observador ( por Pablo Navarro Esteve) pp. 32 y 33.-- Fernández Morales, Angélica. De concreto a conceptual. Relaciones entre arte y arquitectura en el contexto helvético contemporáneo ( por Luis Agustín) pp. 33.—EXPOSICIÓN: Intervenciones cromáticas en los comercios del centro histórico ( por Jorge Llopis verdú) pp. 34 y 35.Seguí De La Riva, J.; Torres Barchino, A.; Gutiérrez Mozo, ME.; Cundari, C.; Chías Navarro, P.; López González, C.; García Valldecabres, J.... (2014). Reseñas. EGA. Revista de Expresión Gráfica Arquitectónica. 19(24):4-35. https://doi.org/10.4995/ega.2014.3268SWORD435192

Multiancestry analysis of the HLA locus in Alzheimer’s and Parkinson’s diseases uncovers a shared adaptive immune response mediated by HLA-DRB1*04 subtypes

Across multiancestry groups, we analyzed Human Leukocyte Antigen (HLA) associations in over 176,000 individuals with Parkinson’s disease (PD) and Alzheimer’s disease (AD) versus controls. We demonstrate that the two diseases share the same protective association at the HLA locus. HLA-specific fine-mapping showed that hierarchical protective effects of HLA-DRB1*04 subtypes best accounted for the association, strongest with HLA-DRB1*04:04 and HLA-DRB1*04:07, and intermediary with HLA-DRB1*04:01 and HLA-DRB1*04:03. The same signal was associated with decreased neurofibrillary tangles in postmortem brains and was associated with reduced tau levels in cerebrospinal fluid and to a lower extent with increased Aβ42. Protective HLA-DRB1*04 subtypes strongly bound the aggregation-prone tau PHF6 sequence, however only when acetylated at a lysine (K311), a common posttranslational modification central to tau aggregation. An HLA-DRB1*04-mediated adaptive immune response decreases PD and AD risks, potentially by acting against tau, offering the possibility of therapeutic avenues

Por una escuela participativa

Memoria Anexo en C-Innov.108 del ejemplar ubicado en el CIDEEste Proyecto pretende alcanzar los siguientes objetivos: Relacionar y coordinar las diversas actuaciones y entidades que inciden en el ámbito educativo conformando la comunidad escolar. Involucrar a los padres y madres en los temas educativos. Atender a las necesidades formativas de los padres y madres. Dinamizar el ambiente cultural del entorno, constituyéndose la escuela en foco motivador. Sentar las bases metodológicas de una educación en valores. Realizar tres actuaciones educativas por curso comunes a todo el centro en el ámbito de la educación en valores. Establecer una comunicación permanente entre los distintos sectores de la comunidad educativa a través de una revista. Utilizar las instalaciones del centro en horario extraescolar. Se realizará una evaluación final del Proyecto y de cada actividad de desarrollo del mismo.Gobierno de Cantabria. Consejería de Educación y JuventudCantabriaES

Dapagliflozin Does Not Modulate Atherosclerosis in Mice with Insulin Resistance

Type 2 diabetes mellitus (T2DM) increases morbimortality in humans via enhanced susceptibility to cardiovascular disease (CVD). Sodium-glucose co-transporter 2 inhibitors (SGLT2i) are drugs designed for T2DM treatment to diminish hyperglycaemia by reducing up to 90% of renal tube glucose reabsorption. Clinical studies also suggest a beneficial action of SGLT2i in heart failure and CVD independent of its hypoglycaemiant effect. In the present study, we explored the effect of SGLT2i dapagliflozin (DAPA) in the metabolism and atherosclerosis in Apoe−/−Irs2+/− mice, which display accelerated atherosclerosis induced by insulin resistance. DAPA treatment of Apoe−/−Irs2+/− mice, which were fed a high-fat, high-cholesterol diet, failed to modify body weight, plasma glucose or lipid. Carbohydrate metabolism characterisation showed no effect of DAPA in the glucose tolerance test (GTT) despite augmented insulin levels during the test. In fact, decreased C-peptide levels in DAPA-treated mice during the GTT suggested impaired insulin release. Consistent with this, DAPA treatment of Apoe−/−Irs2+/− isolated islets displayed lower glucose-stimulated insulin secretion compared with vehicle-treated islets. Moreover, insulin-signalling experiments showed decreased pAKT activation in DAPA-treated adipose tissue indicating impaired insulin signalling in this tissue. No changes were seen in lesion size, vulnerability or content of macrophages, vascular smooth muscle cells, T cells or collagen. DAPA did not affect circulating inflammatory cells or cytokine levels. Hence, this study indicates that DAPA does not protect against atherosclerosis in insulin-resistant mice in hypercholesterolemic conditions